Application and precautions of HEPES in nucleic acid related experiments

HEPES, The full name is 4-hydroxyethyl piperazine ethanesulfonic acid also known as 4- (2-hydroxyethyl) -1-piperazineethanesulonic acid, with a CAS number of 7365-45-9. As a non-volatile zwitterionic chemical buffer, HEPES is widely regarded as one of the excellent universal buffers in the field of biological research. It belongs to the famous Good's biological buffering reagent family and has excellent buffering ability under physiological pH conditions.

HEPES (Cell Culture Grade) products independently developed and produced by Hubei Xindesheng Material Technology Co., Ltd. are widely used in scientific research and production fields such as cell culture, protein purification, molecular biology, etc., with high purity, low endotoxin, and stable process as their core advantages.

1. Basic information

1.1 Chemical and buffering properties

1.2 Buffer performance

The pKa value of HEPES is about 7.5 (25 ° C), and the effective buffering range is pH 6.8-8.2. It exhibits strong buffering ability in the pH range of 6.0-8.5. Especially in the physiological range of pH 7.2-7.4 optimal for cell culture, its buffering capacity is much higher than that of phosphate or carbonate buffer solutions.

Hepes packaging 

As a member of the Good's buffer family, HEPES possesses excellent characteristics such as high water solubility, low cell membrane permeability, stable acid-base dissociation constant, low metal ion chelating ability, high chemical stability, and low absorption in the UV and visible light regions.

2. Core advantages

2.1 A buffer system that does not rely on CO ₂

Traditional cell culture media rely on a bicarbonate/CO ₂ buffer system to maintain pH balance. Once the CO ₂ incubator is opened, there will be severe fluctuations in pH. The buffering capacity of HEPES is independent of CO ₂ concentration. When cells are divided into vials, passaged, or observed microscopically in a biosafety cabinet, the culture medium containing HEPES can maintain pH stability in an open environment for several hours, effectively protecting cells from alkaline environment damage.

2.2 Excellent biocompatibility

At typical working concentrations (10-25 mM), HEPES exhibits good tolerance to most cell lines and has no significant toxic effects on cells. HEPES has chemical and enzymatic stability and cannot penetrate cell membranes, making it highly suitable for addition to cell culture media.

2.3 Excellent temperature stability

Unlike Tris and other buffer solutions, the pKa value of HEPES shows minimal temperature variation (-0.014/° C) and maintains stable buffering capacity within the experimental temperature range of 4 ° C to 37 ° C. This characteristic enables it to maintain the structure and function of the enzyme well even under low temperature conditions.

2.4 Friendly to metal ions

HEPES does not chelate with common metal ions such as Ca ² ⁺ and Mg ² ⁺, and does not interfere with enzyme activity dependent on metal ions, making it suitable for experimental systems related to metal ions.

3.Application scenarios

3.1  Cell culture

HEPES is widely used in mammalian cell culture, tissue engineering, and other research. The commonly used working concentration is 10-25 mM. Compared with inorganic bicarbonate buffer systems, HEPES exhibits superior pH control ability in tissue or organ culture. Especially in scenarios where cells need to be processed outside the CO ₂ incubator for a long time, adding HEPES can significantly enhance the buffering capacity of the culture medium. The conventional culture, long-term incubation, as well as cell crawling and Transwell migration/invasion experiments of sensitive cells (such as primary cells, stem cells, tumor cells) can be maintained by adding HEPES to maintain cell homeostasis.

3.2  Protein purification and extraction

HEPES is a commonly used binding buffer and lysis buffer component in protein research. It has a wide range of applications in biological processes such as purifying antibodies, bioactive substances, proteins, and growth factors.

3.3  Electrophoresis and Molecular Biology

HEPES can be used in gel electrophoresis buffer, DNA/RNA manipulation and other molecular biology experiments.

3.4  Other applications

HEPES can also be used as a biphasic electrolyte solution for isoelectric focusing electrophoresis, a buffer system for observing protein deposition under electron microscopy, and a buffer reagent for quantitative and selective determination of antigen antibody reactions.

HEPES buffer powder

4. New Desheng HEPES Product Specifications and Quality Control

Hubei Xindesheng Material Technology Co., Ltd. was established in 2005 and is a national high-tech enterprise specializing in the research, development, production, and sales of in vitro diagnostic reagents and biological buffering agents. The company has rich technical accumulation in the field of biological buffering agents, with products covering dozens of commonly used and customized types such as TRIS, HEPES, MOPS, etc., with a daily production capacity of up to 5 tons.

Product Features of Desheng

High purity: The purity of cell culture grade HEPES can reach over 99%

Low endotoxin: Strict endotoxin control indicators ensure the smooth progress of cell culture experiments

Powder form: Easy to store and transport for a long time. When using, add deionized water and stir to dissolve. Adjust to the desired pH value with sodium hydroxide. Easy to prepare

Good batch stability: strict quality control system ensures consistent performance of products from different batches

quality assurance

Xindesheng adheres to the business philosophy of "quality first, customer-oriented", and its products are strictly tested according to industry standards. The company is actively building a new factory in Huanggang, committed to creating a domestic base for high-end biological buffering agents, and further enhancing production capacity and supply chain guarantee capabilities.

5.Precautions

5.1 HEPES recommends a working concentration of 10-25 mM. A low concentration is not sufficient to effectively control pH fluctuations, while a high concentration may cause toxicity to certain sensitive cells.

5.2 HEPES can interfere with Folin Ciocalteu protein quantification experiments, but will not affect Biuret protein quantification analysis.

5.3 The product is in non sterile packaging. If used for cell culture, it is recommended to perform sterile filtration before use.

5.4 It is recommended to store at room temperature, away from light, moisture, and in a sealed and dry environment.

Hubei Xindesheng Material Technology Co., Ltd. is committed to providing high-quality biological buffer products for the global life science research and in vitro diagnostic industry. If you need to learn more about our products or obtain samples, please feel free to contact us at any time.