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Sodium Dodecyl Sulfate Polyacrylamide Gel Electrophoresis

With SDS-PAGE analysis, proteins react with SDS by hydrophobic interactions, producing a uniformly negatively charged complex, which is resolved according to its molecular weight. In this case, the relationship between linear migration in the gel and the molecular weight (MW) of the protein is more accurate and by using known molecular weight standards in the same gel, it is possible to estimate the molecular weight of an unknown protein. Resolution of proteins in SDS PAGE is not affected by phosphorylation or isoelectric pH. The electrophoretic pattern of bovine milk is reported in Figure 2.1.4.7, lane 8. In the same figure, lines 2, 3, and 4 represent the casein fraction of bovine milk separated by acidification with acetic acid at pH 4.6. Lines 5 to 7 represent

Urea-PAGE of (1-3) goat caseins, (4-7) sheep caseins, and (8-9) cow caseins

Figure 2.1.4.6 Urea-PAGE of (1-3) goat caseins, (4-7) sheep caseins, and (8-9) cow caseins.

SDS-PAGE of bovine caseins fraction (lanes 1-3), bovine whey proteins (lanes 4-7), skimmed bovine milk (lane 8)

Figure 2.1.4.7 SDS-PAGE of bovine caseins fraction (lanes 1-3), bovine whey proteins (lanes 4-7), skimmed bovine milk (lane 8).

the remaining whey proteins of milk after isoelectric precipitation of caseins. With SDS-PAGE, the different caseins are not resolved, since they have almost the same molecular weight (~23 kDa). Instead, a-lactalbumin and p-lactoblobulin are well resolved since there is a difference of 4 kDa in their MWs.

Magenis, Prudencio, Molognoni, and Daguer (2014) proposed the SDS-PAGE analysis of cheese proteins to assess the compositional authenticity of white nonripened cheeses like Minas Frescal, a typical Brazilian cheese. The analysis aimed to detect the irregular replacement of milk with whey in cheese making. The SDS-PAGE of cheese proteins was performed, followed by image densitometry. p-lactoglobulin (p-LG) was chosen as the adulteration marker, identified by in gel trypsin digestion followed by MALDI-TOF mass spectrometry.

A variant of the SDS-PAGE analysis is the Native PAGE in which denaturing agents such as SDS or Ditrioeritrothyol are not used. This type of electrophoresis is commonly used to evaluate protein to protein interactions (Wittig & Schagger, 2009). Pesic et al. (2011) suggested a native gel electrophoresis of milk for the simultaneous qualitative and quantitative detection of the fraudulent addition of bovine milk to caprine and ovine milk. The test consists of measuring the band intensity of bovine p-lactoglobulins and bovine a-lactalbumin in caprine/bovine milk blends. Linear relationships were established between the band intensity of bovine p-lactoglobulins and the volume percentage of bovine milk added.

 
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