Native gel electrophoresis on acrylamide basis for separating bovine serum proteins

Serum protein electrophoresis (SPEP) of native proteins is a diagnostic tool that helps to differentiate between acute and chronic inflammatory diseases in animals. SPEP is performed in horizontal zone electrophoresis systems with ultra-thin agarose gels or cellulose acetate membranes, or in the capillary zone electrophoresis without carrier material. Biomedical research laboratories more commonly use acrylamide gels as a carrier for protein analysis. As this is not applied for diagnostic SPEP, our study aimed to establish and characterize of an acrylamide-based SPEP with serum samples from cattle. This study showed the best electrophoretic separation of the serum protein fractions by use of concentration gradient gels (2−12% acrylamide/bis-acrylamide in Tris-acetate buffer) and Tris-taurine electrophoresis running buffer. Post-analysis of the protein bands by mass spectrometry identified those proteins contributing to the major SPEP fractions. While some proteins migrated in the expected fractions ( e.g., α1-antiproteinase, complement factor C3), others migrated elsewhere ( e.g., α2-macroglobulin, haptoglobin). In contrast to Tris-acetate/taurine buffer chemistry, Tris-borate proved to be the best buffer condition for separating the serotransferrin-containing fractions, allowing an appropriate differentiation of the individual transferrin serotypes. In summary, our study suggests electrophoresis protocols that can be used for bovine SPEP in acrylamide gels.