Let us getting know the application of TRIS and CAPS in transfer buffer

الوصف

The transfer buffer contains a variety of ingredients that promote protein transfer. The optimization of the buffer is based on the type of transfer system used (wet transfer or semi-dry transfer), the type of gel used, the choice of membrane, and the protein of interest. The transfer buffer must have a strong buffering capacity to keep the conductivity and pH stable during the transfer process. The pH of the buffer must be different from the isoelectric point (pI) of the related protein, otherwise the protein will not transfer.
 
The most common buffer is Tris (Tris) with a pH of 8.**8.5, which is higher than the pI of most proteins. When transferring high molecular weight or basic proteins, higher pH buffers such as CAPS or carbonate can also be used.

The traditional transfer buffer contains a buffer system, methanol, and Tris glycine buffer, and is often used in tank transfer systems. The pH of this buffer is 8.3, which is higher than the isoelectric point of most proteins. The proteins separated on the gel are negatively charged and migrate toward the anode. Since the buffer in the tank is mixed, the ion distribution during the transfer process remains relatively constant.
 
The semi-dry system can be operated with a single or three buffer system. Three buffers are used because transfer is an isotachophoresis process in which the protein moves between the leading ion and the trailing ion. In some cases, the three buffer system can achieve better quantitative transfer. These buffers are:
1. Anode buffer I: 0.3 M Tris, pH *0.4
2. Anode buffer II: *5 mM Tris, pH *0.4
3. Cathode buffer: *5 mM Tris and *0 mM ε-aminocaproic acid, pH 9.4
 
The anode buffer I neutralizes the excess protons generated on the surface of the anode plate. The pH value of anode buffer II and anode buffer I are the same, but the Tris concentration is as low as *5 mM. The cathode buffer contains ε-aminocaproic acid as a trailing ion during the transfer process and is gradually depleted in the cathode buffer as it moves through the gel to the anode. Which single buffer is used in the semi-dry system depends on the specific situation.
 
Although the buffer system defined above is suitable for most protein transfers, it is recommended to use *0 mM CAPS buffer when it is suitable for different applications, especially in protein sequencing applications. Any modification of buffer strength and composition should be done carefully to ensure that the transfer device does not overheat.