Tris HCl usage and treatment: key steps in biochemical experiments

Release time:

2023-12-19


Tris HCl (Trihydroxymethylaminomethane hydrochloride) is a commonly used buffer in biochemical and molecular biology experiments, with a stable pH value and high ion strength. It is an important reagent in experiments such as nucleic acid and protein extraction, electrophoresis, and chromatography. The molecular formula of Tris HCl is C4H11NO3, with a molecular weight of 157.6. It is a compound composed of trimethylaminomethane (Tris) and hydrochloric acid (HCl), with a white crystalline powder appearance. It is easily soluble in water and insoluble in organic solvents, exhibiting good buffering performance. Below, we will delve into the usage and processing methods of Tris HCl in experiments, as well as some aspects that need to be noted.

Buffering mechanism

Tris HCl can maintain the acid-base balance of the solution, and its effect mainly depends on the amino and carboxyl groups in its molecules. Amino groups can react with acidic substances to maintain the alkalinity of the solution, while carboxyl groups can react with alkaline substances to maintain the acidity of the solution. Therefore, Tris HCl can effectively prevent pH fluctuations in the solution and protect the stability of biomolecules. In addition, Tris HCl can also improve the solubility of the sample and promote the separation and purification of biomolecules.

Tris HCl usage method

Tris HCl is usually supplied in powder form, and when used, Tris HCl powder is dissolved in distilled water or deionized water according to the required buffer concentration and volume. The recommended buffer concentration is generally 0.05-0.1M, with a pH value of 7.5-8.5. Samples treated with Tris HCl should be stirred evenly. If it is necessary to adjust the pH value, slowly add acid or alkali and continuously stir.

Tris HCl usage and handling precautions

1. Tris HCl has a high ion strength and is not suitable for experiments that require low ion strength.

2. When preparing Tris HCl buffer, clean containers and deionized water should be used. If unclean containers or water containing impurities are used, it will affect the pH and purity of the buffer.

3. When using Tris HCl, an appropriate pH range should be selected according to experimental requirements. The suitable pH for Tris HCl at 25 ° C is around 7.5, which can be adjusted by temperature and the concentration of Tris and HCl.

4. The buffering effect of Tris HCl is sensitive to temperature, and under high temperature conditions, its buffering effect will decrease. In some experiments that require high temperature treatment, careful selection or additional measures should be taken.

5. Tris HCl should be stored in a dry, cool environment, away from light, and kept away from moisture to prevent degradation.

Processed Tris HCl application

1. Protein extraction: In the process of protein purification, Tris HCl is used as the main component of cell lysis buffer, whose buffering ability enables the lysis buffer to maintain the structure and activity of proteins while maintaining an alkaline environment.

2. Electrophoresis buffer: Tris HCl is also often used as the electrophoresis buffer in SDS-PAGE (polyacrylamide gel electrophoresis) to separate proteins. Its pH buffer ensures the stability of proteins during electrophoresis.

3. DNA/RNA extraction: Tris HCl is widely used in DNA and RNA extraction, ensuring the stability of nucleic acids during the extraction process and preventing degradation and damage.

4. PCR buffer: In polymerase chain reaction (PCR), Tris HCl is often used as a component of the PCR buffer to maintain the pH of the reaction system and promote the activity of DNA polymerase.

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