Buffer for electrophoresis

Tris-acetic acid buffers have been proposed instead of Tris-hydrochloric acid buffers for preparative starch gel electrophoresis because the anodic solution does not drop to such low pH values (Pilz and Johann, 1966). Examples for use at 20°C are pH 7.3, four litres of 0.4m Tris plus 750 ml 2m acetic acid pH 8.6, 4.5 litres 0.4m Tris plus 280 ml 2m acetic acid. A recent paper describes buffers for the electrophoresis of enzymes on polyacrylamide gel (Orr et al., 1972). 

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Table 8.1 Continuous Buffers for Electrophoresis of Native Proteins...

The buffer system is a combination of buffer for electrophoresis and eluent for the particular chromatographic mode being employed. Figure 5.14 shows the separation of a group of neutral molecules using a capillary packed with a reversed-phase material.38 The buffer was a mixture of 4 mM sodium tetraborate (pH 9.1) and acetonitrile (20 80, v/v). The separation was compared with a micro-HPLC separation in which the same capillary was used but the eluent was pressure driven. As can be seen in Figure 5.14, sharper peaks were obtained with the EOF-driven system. 

A buffer for electrophoresis is usually prepared at room temperature by adding ingredients and measuring the final pH. When this buffer is used for CE at a different temperature, a different pH might result. For example, when a buffer consists of Tris-acetate at pH 5, Tris is fully charged and the temperature dependence of pK of acetic acid is the one involved. This value is small so that the pH of the buffer is virtually inde-... 

Dissolve the lyophilized proteolytic digests in at least 6 p pH 1.9 buffer or pH 4. 72 buffer (for electrophoresis at pH 1.9 and pH 4.72, respectively) or deionized water (for electrophoresis at other pH s), using a vortex mixer (the final volume depends on the total number of counts per minute in the sample). Centrifuge the sample briefly in a microcentrifuge to ensure that any residual particulate matter is not applied to the plate. 

Table 3.11 Constant ionic strength buffers for electrophoresis (2°C) ... 

X TBE buffer or1X TAE buffer Make 0.5X TBE buffer or 1X TAE buffer for electrophoresis and agarose gels. 

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