HHBS (Hank's Buffer with Hepes) Preparation and Recipe

HHBS is a general purpose buffer that combines the stability of Hank's buffer with the good buffering capacity of HEPES buffering solution (one of Good's buffers). HHBS is used extensively for cell culture, but is sensitive to light.

Available for purchase here.

To prepare

L of HHBS (Hank's Buffer with Hepes):

Change the value in the textbox above to scale the recipe volume

Table 1. Required components

Component	Amount	Concentration
Calcium Chloride (mw: 110.98 g / mol)	#	#
Magnesium Chloride Hexahydrate (mw: 203.3 g / mol)	#	#
Magesium Sulfate Heptahydrate (mw: 246.47 g / mol)	#	#
Potassium Chloride (mw: 74.55 g / mol)	#	#
Potassium Phosphate Monobasic (mw: 136.09 g / mol)	#	#
Sodium bicarbonate (mw: 84.01 g / mol)	#	#
Sodium chloride (mw: 58.44 g / mol)	#	#
Sodium Phosphate Dibasic (mw: 141.96 g / mol)	#	#
D-Glucose (Dextrose) (mw: 180.16 g / mol)	#	#
Hepes (mw: 238.3 g / mol)	#	#

Prepare 800 mL of distilled water in a suitable container.
Add # of Calcium Chloride to the solution
Add # of Magnesium Chloride Hexahydrate to the solution
Add # of Magesium Sulfate Heptahydrate to the solution
Add # of Potassium Chloride to the solution
Add # of Potassium Phosphate Monobasic to the solution
Add # of Sodium bicarbonate to the solution
Add # of Sodium chloride to the solution
Add # of Sodium Phosphate Dibasic to the solution
Add # of D-Glucose (Dextrose) to the solution
Add # of Hepes to the solution
Add distilled water until the volume is 1 L.

References

This online resource may be cited as follows

MLA	"Quest Calculate™ HHBS (Hank's Buffer with Hepes) Preparation and Recipe." AAT Bioquest, Inc., 27 Oct. 2025, https://www.aatbio.com/resources/buffer-preparations-and-recipes/hhbs-hanks-buffer-with-hepes.
APA	AAT Bioquest, Inc. (2025, October 27). Quest Calculate™ HHBS (Hank's Buffer with Hepes) Preparation and Recipe. AAT Bioquest. https://www.aatbio.com/resources/buffer-preparations-and-recipes/hhbs-hanks-buffer-with-hepes.
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This online resource has been cited in 1 publications, including

Identification of Autophagy as a Functional Target Suitable for the Pharmacological Treatment of Mitochondrial Membrane Protein-Associated Neurodegeneration (MPAN) In Vitro

Authors:

Zanuttigh, Enrica and Derderian, Kevork and G{\"u}ra, Miriam A and Geerlof, Arie and Di Meo, Ivano and Cavestro, Chiara and Hempfling, Stefan and Ortiz-Collazos, Stephanie and Mauthe, Mario and Kmie{\'c}, Tomasz and others,

Journal:

Pharmaceutics (2023): 267