In all kinds of cell, protein, and DNA experiments, once the components leave the natural biological environment, they will be seriously affected. To provide them with a good natural environment, buffer must be used, TRIS, phosphate, HEPES (4-hydroxyethylpiperazineethanesulfonic acid), etc...... then how to choose a most suitable buffer?

Let's take a look at the commonly used biological buffers, and their characteristics:

1. Phosphate buffer

Phosphate buffer is the most widely used buffer. Due to its secondary dissociation, the buffer has a wide pH range and can be configured with acidic, basic and neutral buffers of various pH values:

•  Preparation of acidic buffer can directly use NaH₂PO₄ or KH₂PO₄, the pH range is 1~5;
• Alkaline buffer can directly use Na₂HPO₄ or K₂HPO₄, pH range is 9~12;
•  Neutral buffer contains equal concentrations of NaH₂PO₄ and Na₂HPO₄ or equal concentrations of KH₂PO₄ and K₂HPO₄ solutions at pH 5.5~8.5.

2. TRIS buffer

Tris buffer is widely used in biochemical research. It is a weak base and usually used in the "neutral" range. Tris-HCl buffer: pH=7.5~8.5; Tris-phosphate Buffer: pH = 5.0 to 9.0.

In addition to Tris-HCl, TRIS has a variety of derivatization buffers:

• TBS=Tris-HCl+ NaCl+KCl, commonly used to clean immunostained tissues or Western blotting membranes in Western Blotting;
• TBST=Tris-HCl+NaCl+tween20, a membrane buffer commonly used in Western Blotting;
• TE=Tris-HCl+EDTA, which is protective against DNA bases and is often used for DNA stabilization and storage;
• TAE=Tris base+acetic acid+EDTA, which is a buffer system widely used in short-segment DNA electrophoresis;
• TBE=Trisbase+boric acid+EDTA, suitable for long-term DNA electrophoresis, and has better separation effect on smaller fragments.

3. Glycine buffer

Glycine buffer has a wide pH range and a wide range of applications. The pH range is 2.0~11.0. The most common system and pH range are shown in the following table:

4. HEPES buffer

HEPES is a non-ionic amphoteric buffer with good buffering capacity in the range of pH 7.2~7.4. It is commonly used in biochemical diagnostic kits, DNA/RNA extraction kits and PCR diagnostic kits.

Table 1. Advantages and disadvantages of commonly used buffers

General principles for selecting buffers:

(1) The most important thing is to determine the pH value required by your system.

(2) Select the buffer system, the buffering capacity will decrease significantly when the pKa changes more than one pH unit;

(3) The buffer system should not affect the activity of the protein or the stability of the DNA, and avoid interference from other ions;

(4) Finally, we must also consider the effect of temperature: temperature not only affects the buffer, but also affects the cells. Most animal cells have a pH of 7.0 to 7.5 at 37 °C, and when the temperature falls to 0 °C, it can reach 8.0.

Edited by Suzhou Yacoo Science Co., Ltd.