Recipes of HEPES Buffer

4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid (HEPES) is a hydrogen ion buffer which has a good buffering capacity in the pH range of 6.8~8.2 and can control a constant pH for a long time. It can be used in a wide range of biochemical reactions, such as, in the research of organelles and highly denatured pH-sensitive proteins and enzymes, as well as biochemical diagnostic kit, DNA/RNA extraction kits and PCR diagnostic kit. Below we will list the common used recipes.


(1) HEPES buffer

Reagent	Amount to add (for 1 L)	Final concentration
NaCl	6.72 g	115 mM
CaCl2	133 mg	1.2 mM
MgCl2	114 mg	1.2 mM
K2HPO4	418 mg	2.4 mM
HEPES	4.77 g	20 mM
H2O	to 1 L

Adjust the pH to 7.4 with HCl or NaOH. HEPES buffer can be stored refrigerated for several weeks.
(2) HEPES Binding Buffer
10-mM HEPES, pH 7.4; 150 mM NaCl; 5 mM KCl; 1 mM MgCl2; 1.8 mM CaCl2.

• Annexin V-FITC in HEPES binding buffer

Annexin V-FITC can be obtained from different companies (e.g., Clontech, Boehringer Mannheim, BioWhittaker). We recommend titrating it to determine optimal concentrations.

• PI in HEPES binding buffer

100 µg/mL propidium iodide, Dissolve propidium iodide in HEPES binding buffer.
(3) HEPES-KRH Buffer

Reagent	Final concentration
NaCl	116 mM
KCl	4 mM
MgCl2	1 mM
CaCl2	1.8 mM
Glucose	25 mM
HEPES acid	10 mM

Adjust pH to 7.4.

(4) HEPES-buffered saline buffer

Reagent	Final concentration
NaCl	150 mM
Tween-20	0.005% (v/v)
EDTA	3 mM
HEPES pH (7.4)	10 mM

(5) HEPES-Buffered Saline (HBS) Precipitation Buffer (2×)
50 mM HEPES, 280 mM NaCl. Adjust the pH to 7.10 (±0.05) with NaOH. Bring the final volume to 1 L in ddH2O. Filter-sterilize.
(6) HEPES Immunoprecipitation Buffer

Reagent	Final concentration
NaCl	10 mM
Na-β-glycerophosphate	20 mM
EDTA	1 mM
HEPES pH (7.5)	50 mM
Na3VO4	0.1 mM
NaF	50 mM
PMSF	1 mM
DTT	1 mM
NP-40	0.1%
Protease inhibitor cocktail

Edited by Suzhou Yacoo Science Co., Ltd.