单组分溶液配制

互联网2013-09-05

5838

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 ".

See also:

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).

<center><font>About the recalculation of recipes for the arbitrary volumes:<br /> <br /> </font> </center>

Organic substances.

pK_a and temperature dependence of pH for common buffers.

Buffer	Mw	pK_a 20^o C	Working range	delta pK_a per 10^o C
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.undefined	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

ATP

C₁₀ H₁₄ N₅ O₁₃ P₃ Na₂ Mw=551.1 g/M; (store at -20^o C).

	Conc.	Stock	1ml
ATP	0.1M	551.1g/M	55.1mg
H₂ O		mQ

0.1M ATP (pH 7.2): 5.71% (w/w) ATP, 84.86% (w/w) H₂ O, 9.43% (w/w) 2M Tris base.

adjust to pH7.5 by 2M NaOH (~70-80µl);
prepare about 700µl, dilute 2000 times (it is the final dilution in the spectrofotometric cell), check the optical density:
C[M]=A₂₅₉ /15.4x10³ =A_{259 1:2000} x0.130
adjust the final volume.

Betaine

monohydrate C₅ H₁₁ NO₂ xH₂ O; Mw=135.2g/M; (store at 4^o C).

	Conc.	Stock	%(w/w)	100ml	200ml	700ml
Betaine	135.2g/M	5M	63.00	67.6g	135.2g	473.2g
H₂ O		mQ	37.00	39.74ml	79.49ml	278.2ml

p~1.073g/ml

Cresol red (Na)

50mM (store at +4, -20^o C):

	Conc.	Stock	1ml	50ml
Cresol red (Na)	50mM	404.4g/M	20.2mg	1.01g
H₂ O		mQ

Cresol Red (Na salt) is a very convenient dye. At a concentration of about 0.2mM it is compatible with restriction digestion, PCR, sequencing. It may be used as marker for electrophoresis;

the color of the dye is pH-dependent (red, if pH>7.5; yellow, if pH <7.0). It is possible to use Cresol Red as pH-indicator (i) for denaturation of double-stranded DNA in sequencing, (ii) for silica-purification of DNA from agarose gel.

DTT

C₄ H₁₀ O₂ S₂ Mw=154.2g/M; (store at -20^o C).

	Conc.	Stock	%(w/w)	1ml	5ml	10ml	20ml
DTT	154.2g/M	1M	14.55	0.155g	0.773g	1.55g	3.09g
AcNa, pH 5.2		10mM	85.45	905 µl	4.53ml	9.05ml	18ml

p~1.06g/ml

	Conc.	Stock	1ml	5ml	10ml	20ml
DTT	154.2g/M	2.2M	0.339g	1.696g	3.394g	6.787g
AcNa, pH 5.2		10mM

filter sterililize;

no DEPC treatment.

Preparation of dNTP's.

The quick protocol for ~100mM stock.

dilute all four dNTP's (250mg of each) in 3.676ml of H₂ O;
+ 424µl 5M NaOH;
check the pH: ~0.5µl on pH-paper.

Accurate protocol for 100mM stocks preparation.

add the necessary quantity (see table) of H₂ O 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	C₁₀ H₁₄ N₅ O₁₂ P₃ Na₂ x3H₂ O	589.2	3.24ml	850 µl	4.24ml
dGTP	C₁₀ H₁₃ N₅ O₁₃ P₃ Na₃ x2H₂ O	609.2	3.55ml	400 µl	4.10ml
dCTP	C₉ H₁₃ N₃ O₁₃ P₃ Na₃ x2H₂ O	569.1	3.44ml	800 µl	4.39ml
dTTP	C₁₀ H₁₄ N₂ O₁₄ P₃ Na₃ x2H₂ O	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_1:5000 xA_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.

EDTA

C₁₀ H₁₄ O₈ N₂ Na₂ x2H₂ O; Mw=372.3g/M; pH = 8.0 (store at 4^o C).

	Conc.	Stock	%(w/w)	50 ml	100 ml	150 ml	250 ml
EDTA	0.5 M	372.3g/M	16.98	9.31g	18.62g	27.92g	46.55g
NaOH	~0.5 M	40g/M	1.82	1.014g	2.028g	3.042g	5.07g
H₂ O		mQ	81.19	44.48ml	88.95ml	133.4ml	222.4ml

	Conc.	Stock	10 ml	50 ml	100 ml	150 ml	250 ml
EDTA	0.5 M	372.3g/M	1.86g	9.31g	18.62g	27.92g	46.55g
NaOH	~0.5 M	10M	507µl 0.674g	2.535ml 3.37g	5.07ml 6.74g	7.61ml 10.11g	12.68ml 16.85g
H₂ O		mQ	8.42ml	42.12ml	84.24ml	126.4ml	210.6ml

p~1.096g/ml

EDTA is not soluble at acidic pH; it is necessary to add alkali gradually and to control pH;

do not treat by DEPC.

EtBr

Ethidium bromide, C₂₁ H₂₀ N₃ Br; Mw=394.3g/M; (store at NT in the dark);

	Conc.	5ml	10ml	50ml
EtBr	10mg/ml	50mg	100mg	500mg

soluble in H₂ O, EtOH, chloroform;

concerning carcinogenic properties of EtBr. The only data that we found in the literature is that in mutagenic test (on bacteria) 90µg of EtBr gave the same results as the smoke concentrate from one cigarette.

Gelatin

(store at 4^o C).

	Conc.	Stock	10ml	50ml	100ml
Gelatin	2%	solid	0.2g	1.0g	2.0g
H₂ O		mQ

sterilize by autoclaving.

Glucose

C₆ H₁₂ O₆ xH₂ O, Mw=198.17g/M (store at 4^o C)

	Conc.	Stock	%(w/w)	50ml	100ml	250ml
Glucose	2M	198.17g/M	34.972	19.82g	39.63g	99.09g
H₂ O		mQ	65.028	36.85ml	73.70ml	184.24ml

	1000ml/
198.17g/M	1M	1.5M	2M
Glucose [g]	198.17	297.255	396.34
H₂ O [ml]	868.23	802.745	736.96
%(w/w)	18.583	27.023	34.972
H₂ O %(w/w)	81.417	72.977	65.028
p (g/ml) 20^o C	1.0664	1.1	1.1333

Guanidine HCl (GuHCl)

CH₅ N₃ xHCl, Mw=95.53g/M

	Molarity; 1000ml /
1	2	3	4	5	6	7	8
GuHCl	95.53g	191.06g	286.59g	382.12g	477.65g	573.18g	668.71g	764.24g
H₂ O (mQ)	924.2ml	854ml	783.7ml	711.7ml	639.8ml	567.2ml	494.3ml	420.7ml
GuHCl %(w/w)	9.37	18.28	26.78	34.93	42.75	50.26	57.50	64.50
H₂ O %(w/w)	90.63	81.72	73.22	65.07	57.25	49.74	42.50	35.50
p (g/ml)	1.020	1.045	1.070	1.094	1.117	1.140	1.163	1.185

solubility: at 25^o C - 8.54M, 5^o C - > 8M;
A₂₆₀ (6M in H₂ O)<0.03;
it is possible to take the "partial density of GuHCl" as 0.763 in calculations of solutions.

HEPES

	Conc.	Stock	%(w/w)	1L
C₈ H₁₈ N₂ O₄ S	1M	238.3g/M	22.40	238.3g
H₂ O		mQ	77.60	825.7ml

p=1.064

HEPES, 1M, 1L

Desired pH	KOH, 5M [1000ml]

5.25	0ml	0ml
5.35	0.5ml
5.75	3.5ml
6.03	7ml
6.24	12ml
6.59	22ml
6.71	32ml
6.88	45ml
7.00	50ml
7.10	60ml
7.25	80ml
7.37	92.5ml

Imidazol

C₃ H₄ N₂ , (store at 4^o C):

	Conc.	Stock	50ml	100ml
Imidazole	2M	68.1g/M	6.81g	13.62g
H₂ O		mQ

Paraformaldehyde

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

1. mix in the screw-cap tube:

PFA (solid) = 0.37g,
H₂ O = 1.0ml
NaOH (1N) = 14µl;

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

PEG

H(OCH₂ CH₂ )_n OH; (store at 4^o C).

	Conc.	%(w/w)	10ml	50ml	100ml	150ml	200ml
PEG6000	40%	37.21	4.0g	20g	40g	60g	80g
H₂ O	mQ	62.79	6.75g	33.75g	67.5g	101.25g	135.0g

p=1.075.

PMSF

(store at -20^o C)

	Conc.	Stock	20ml
C₇ H₇ FO₂ S	100mM	174.2g/M	0.348g
Isopropanol			20ml

Retinoic acid

all trans-Retinoic acid, Tretinoin, light-sensitive, (store at -20^o C):

	Conc.	Stock	16.6ml
C₂₀ H₂₈ O₂	10mM	300.4g/M	50mg
EtOH		> 96%	16.6ml

stock solution is 10mM, working solution is freshly prepared 1mM in EtOH (it would be better to add pure EtOH to the control cells).

Sucrose

C₁₂ H₂₂ O₁₁ , Mw=342.30g/M; 20^o C.

Densities and refraction indexes

of sucrose solutions.

	Conc.	Stock	%(w/w)	50ml	100ml	250ml
Sucrose	1M	342.30g/M	30.330	17.115g	34.23g	85.58g
H₂ O		mQ	69.670	39.315ml	78.63ml	196.58ml

	1000ml/
342.30g/M	1M	2M	2.5M
Sucrose [g]	342.3	684.6	855.75
H₂ O [ml]	786.3	570.4	460.35
%(w/w)	30.330	54.550	65.022
H₂ O %(w/w)	69.670	45.450	34.978
p (g/ml) 18^o C	1.1286	1.2550	1.3161

Tris Cl

C₄ H₁₁ O₃ N; Mw=121.1g/M; (store at 4^o C).

	Conc.	Stock	50 ml	100 ml	150ml	200ml
Tris-base	1M	121.1g/M	6.06	12.11g	18.17g	24.22g
H₂ O to the final weight		mQ	52.03g	104.06g	156.09g	208.12g

1M: p=1.0406

2M Tris base: 22.90%(w/w) Tris base, 77.10%(w/w) H₂ O; p=1.058

do not treat by DEPC;
sterilize by autoclaving;
pH of Tris-buffers is dependent from concentration. If to take 50mM solution as the original:
pH(500mM) => + 0.05
pH(5mM) => - 0.05
pH drops on 0.028 when the temperature rise on 1^o C.

Temperature dependence of pH for Tris Cl 50mM.

pH at	g/50ml 1M or g/liter for 0.05 M
5^o C	25^o C	37^o C	Tris HCl	Tris Base	H₂ O	Tris HCl	Tris Base	H₂ O
7.55	7.00	6.70	7.28	0.47	44.28
7.66	7.10	6.80	7.13	0.57	44.33
7.76	7.20	6.91	7.02	0.67	44.34
7.89	7.30	7.02	6.85	0.80	44.38
7.97	7.40	7.12	6.61	0.97	44.45
8.07	7.50	7.22	6.35	1.18	44.50
8.18	7.60	7.30	6.06	1.39	44.58
8.26	7.70	7.40	5.72	1.66	44.65
8.37	7.80	7.52	5.32	1.97	44.74
8.48	7.90	7.62	4.88	2.30	44.85
8.58	8.00	7.71	4.44	2.65	44.94
8.68	8.10	7.80	4.02	2.97	45.04
8.78	8.20	7.91	3.54	3.34	45.15
8.88	8.30	8.01	3.07	3.70	45.26
8.98	8.40	8.10	2.64	4.03	45.36
9.09	8.50	8.22	2.21	4.36	45.46
9.18	8.60	8.31	1.83	4.65	45.55
9.28	8.70	8.42	1.50	4.90	45.63
9.36	8.80	8.51	1.23	5.13	45.67
9.47	8.90	8.62	0.96	5.32	45.75
9.56	9.00	8.70	0.76	5.47	45.80
9.67	9.10	8.79	0.69	5.53	45.81

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

Tricine

C₆ H₁₃ NO₅ , Mw=179.2g/M;(store at 4^o C).

	Conc.	Stock	50ml
Tricine	1M	179.2g/M	8.96g
H₂ O		mQ

For 50ml:

Desired pH	5N KOH [50ml]
8.30	7.0ml
8.38	7.5ml
8.47	8.0ml
8.5	8.15ml
8.6	8.5ml
8.68	9.0ml
8.78	9.5ml
8.90	10.0ml

Triethanolamine

1M (store at 4^o C):

	Conc.	Stock	%(w/w)	50ml
Triethanolamine	1M	149.19g/M	14.69	7.46g 6.66ml
H₂ O		mQ	85.31	43.34ml

Urea

CH₄ N₂ O, Mw=60.06g/M;

	Molarity; 1000ml /
1	2	3	4	5	6	7	8	9	10
CH₄ N₂ O [g]	60.06	120.12	180.18	240.24	300.30	360.36	420.42	480.48	540.54	600.60
H₂ O [ml]	950.6	905.8	861.3	817.0	771.6	726.7	681.2	635.7	590.0	544.1
p (g/ml)	1.011	1.026	1.041	1.057	1.072	1.087	1.102	1.116	1.131	1.145
CH₄ N₂ O %(w/w)	5.94	11.71	17.30	22.72	28.02	33.15	38.16	43.05	47.81	52.47
H₂ O %(w/w)	94.06	88.29	82.70	77.28	71.98	66.85	61.84	59.95	52.19	47.53

solubility: at 25^o C: 10.49M, 5^o C: ~8M;
A₂₆₀ (6M in H₂ O)<0.06;
it is possible to take the "partial density of urea" as 0.763 in calculation of solutions.

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	NH₄ OH	35.0	28%	14.8	251	0.898	67.6
Acetic acid, glacial	CH₃ COOH	60.05	99.5%	17.4	1045	1.05	57.5
Acetic acid	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
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
60%	9.2	923	1.54	108.7
Phosphoric acid	H₃ PO₄	98.0	85%	18.1	1445	1.71	55.2
Sulfuric acid	H₂ SO₄	98.1	96%	18.0	1766	1.84	55.6

NaOH

Mw=40g/M; (store at NT).

	Conc.	Stock	%(w/w)	50ml	200ml	300ml
NaOH	10M	40g/M	30.1	20g	80g	120g
H₂ O		mQ	69.90	46.45ml	185.8ml	278.7ml

10M: 30.10%; p=1.329.

	Conc.	Stock	%(w/w)	50ml	150ml	200ml
NaOH	1M	10M	12.87	5ml 6.645g	15ml 19.94g	20ml 26.58g
H₂ O		mQ	87.13	45ml	135ml	180ml

it is better to use plastic bottles for storage, because the alkali slightly solubilize the glass.

KOH

Mw=56.11g/M; (store at NT).

	Conc.	Stock	%(w/w)	10ml	50ml	100ml	150ml
KOH	5M	56.11g/M	23.05	2.806g	14.03g	28.06g	42.08g
H₂ O		mQ	76.95	9.363g	46.81g	93.63g	140.4g

5M: 23.06%; p=1.217.

it is better to use plastic bottles for storage, because the alkali slightly solubilize the glass.

TCA

CCl₃ CO₂ H Mw=163.39g/M,(store at NT, in the dark, under the fume hood).

	Conc.	%(w/w)	5ml	10ml	25ml	50ml
TCA	100% (w/v)	68.78	5.0g	10.0g	25.0g	50.0g
H₂ O	mQ	31.22	2.27ml	4.54ml	11.35ml	22.7ml

Detergents.

Detergent	T_melt	Mw [Da]	CMC
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
C₁₆ TAB	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	-	-
C₁₂ E₈	-	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

CMC - critical micelle concentration.

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
C₁₆ TAB	+	+	+	-	-	-	+	-	+	-	+	-
Amphoteric
LysoPC	+/-	-	-	-	-	-	-	+	+/-	+	+	-
CHAPS	-	+	-	-	-	-	-	+	+	-	+	-
Zwittergent 3-14	+/-	+/-	-	-	-	-	-	+	+	-	+	-
Nonionic
Octyl glucoside	-	+	-	-	-	-	-	+	-	+	+	-
Digitonin	-	-	-	-	-	-	-	+	+	-
C₁₂ E₈	-	-	-	+/-	-	-	-	+	-	+	-	+
Lubrol PX	-	-	-	+/-	-	+/-	-	-	-	+	-	+
Triton X-100	-	-	-	+/-	+	+/-	-	-	-	+	-	+
Nonident P-40	-	-	-	+/-	+	+/-	-	-	-	+	-	+
Tween 80	-	-	-	+/-	-	+/-	-	-	-	+	-	+

C₁₆ TAB - hexadecyl trimethylammonium bromide;
LysoPC - lysophosphatidylcholine.
~undefined Sodium cholate and sodium deoxycholate are unsoluble at <pH 7.5 or at ionic strength greater then 0.1%. SDS may precipitate below 20^o C;
~undefined_Ionic_detergents_may_induce_problems_with_electrophoresis_and_isoelectric_focusing~J~Kbr_~H~M~undefined*_It_is_possible_to_remove_by_dialysis~J~Kbr_~H~M~undefined**_Phenol_~Fcontaining_detergents_~Afor_example_Triton_X~F100_and_NP~F40~B_precipitate_during_Folin_protein_assay_~Abut_do_not_interfere_with_Bradford_protein_assay~B._~Kbr_~H~M~1~Kbr_~H~M~1~K~Hfont~M~1~1~Ka_name~L~4a29~4~M~K~Ha~M~1~1~Kp_class~L~4h3~4~M~Kfont~MN~FLauroylsarcosine_Na~K~Hfont~M~1~K~Hp~M~1~1~Kp~M~Kfont~M~Astore_at_NT~B.~K~Hfont~M~1~K~Hp~M~1~1~1~1~1~Ktable~M~Ktbody~M~Ktr~M~Ktd~M~1~1~1~1~1~1~1~1~1~1~1~1~1~1~1~1~1~1~1~1~1~1~1~1~1~1~1~1~1~1~1~1~1~1~1~Ktable~M~Ktbody~M~Ktr~M~Ktd~M~1~Kfont~M~K~Hfont~M~1~K~Htd~M~1~Ktd~M~Kfont~MConc.~K~Hfont~M~1~K~Htd~M~1~Ktd~M~Kfont~MStock~K~Hfont~M~1~K~Htd~M~1~Ktd~M~Kfont~M~7~Aw~Hw~B~K~Hfont~M~1~K~Htd~M~1~Ktd~M~Kfont~M1ml~K~Hfont~M~1~K~Htd~M~1~Ktd~M~Kfont~M3ml~K~Hfont~M~1~K~Htd~M~1~Ktd~M~Kfont~M5ml~K~Hfont~M~1~K~Htd~M~1~Ktd~M~Kfont~M15ml~K~Hfont~M~1~K~Htd~M~1~Ktd~M~Kfont~M50ml~K~Hfont~M~1~K~Htd~M~1~Ktd~M~1~Kfont~M~K~Hfont~M~1~K~Htd~M~1~K~Htr~M~1~Ktr~M~Ktd~M~Kfont~MSarc~K~Hfont~M~1~K~Htd~M~1~Ktd~M~Kfont~M10~7~K~Hfont~M~1~K~Htd~M~1~Ktd~M~Kfont~M30~7~K~Hfont~M~1~K~Htd~M~1~Ktd~M~Kfont~M33.93~K~Hfont~M~1~K~Htd~M~1~Ktd~M~Kfont~M~Kfont~M~Ku~M0.333ml~K~Hu~M~1~Kbr_~H~M~10.342g~K~Hfont~M~1~K~Hfont~M~1~K~Htd~M~1~Ktd~M~Kfont~M~Kfont~M~Ku~M1.00ml~K~Hu~M~1~Kbr_~H~M~11.027g~K~Hfont~M~1~K~Hfont~M~1~K~Htd~M~1~Ktd~M~Kfont~M~Kfont~M~Ku~M1.67ml~K~Hu~M~1~Kbr_~H~M~11.712g~K~Hfont~M~1~K~Hfont~M~1~K~Htd~M~1~Ktd~M~Kfont~M~Kfont~M~Ku~M5.00ml~K~Hu~M~1~Kbr_~H~M~15.14g~K~Hfont~M~1~K~Hfont~M~1~K~Htd~M~1~Ktd~M~Kfont~M~Kfont~M~Ku~M16.67ml~K~Hu~M~1~Kbr_~H~M~117.12g~K~Hfont~M~1~K~Hfont~M~1~K~Htd~M~1~Ktd~M~1~Kbr_~H~M~1~1~Kfont~M~K~Hfont~M~1~K~Htd~M~1~K~Htr~M~1~Ktr~M~Ktd~M~Kfont~MH~Ksub~M2~K~Hsub~M~1O~K~Hfont~M~1~K~Htd~M~1~Ktd~M~Kfont~M

66.07

0.667ml

2.00ml

3.33ml

10.00ml

33.33ml

p=1.017.

SDS

Sodium dodecyl sulfate, sodium lauryl sulfate; [CH₃ (CH₂ )₁₀ CH₂ SO₄ ]Na; Mw=288.4g/M; (store at NT).

	Conc.	Stock	%(w/w)	100 ml	250 ml	300 ml	400 ml
SDS	10%(w/v)	solid	9.82	10g	25g	30g	40g
H₂ O		mQ	90.18	91.8 ml	229.5 ml	275.4 ml	367.2 ml

p = 1.018

weight under the fume hood (and better to wear the mask);

it is necessary to heat to 60-80^o C. to facilitate solubilization. Check pH. If it differs from neutral (~7.2-7.5) dramatically - adjust by diluted alkali / acid.

Organic solvents.

Phenol

C₆ H₅ OH; Mw=94.1g/M
p=1.054, t_m = 43, t_b = 182, pKa=10.0
solubility: 6.8¹⁶ H₂ O, unlimited ⁶⁶ H₂ O; unlimited EtOH

Preparation of "acidic" and "neutral" phenol.

a) from the distillation:

distill phenol under H₂ O;
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 H₂ O = 0.02% relative to phenol phase);
---- on this step you obtained the "acidic phenol". Store at -20^o C (~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^o C (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.

______________
~undefined For RNA extraction it is better to saturate (just add) "acidic phenol" by the following buffer: (store at 4^o C):

	Conc.	Stock	100ml
AcONa, pH 5.1	50mM	3M	1.67ml
EDTA	10mM	0.5M	2.0ml
H₂ O		mQ	96.0ml