Two-Dimensional Electrophoresis

Different combinations of the above-mentioned techniques are possible in a double run on the same sample; the most frequent is IEF in the first dimension followed by an SDS-PAGE gradient gel. For specific purposes, such as the study of phosphoproteome and natural variants, the Urea-PAGE could be used instead of SDS-PAGE as first dimension (Mamone et al., 2003). The 2-DGE (IEF/SDS-PAGE) of whole proteins of bovine milk are shown in Figure 2.1.4.9. The identification of spots is in accordance with the study by Holland, Deeth, and Alewood (2004). Thanks to the high resolution, this technique is preferred when used to separate protein before their identification by trypsin digestion followed by mass spectrometry. The 2D-EF is highly complicated, time consuming, and expensive, and its use requires qualified expertise. Therefore, it is used only for research purposes.

IEF of casein fractions after chymosin action, using a pH range 3-10. Lanes 1,3 and 4 caprine, 2 bovine, 5 ovine curd

Figure 2.1.4.8 IEF of casein fractions after chymosin action, using a pH range 3-10. Lanes 1,3 and 4 caprine, 2 bovine, 5 ovine curd.

DGE of bovine milk, protein spots are identified according to (Holland et al., 2004)

Figure 2.1.4.9 2-DGE of bovine milk, protein spots are identified according to (Holland et al., 2004).

 
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