The Optimized Conditions of Two Dimensional Polyacrylamide Gel Electrophoresis for Serum Proteomics

Serum proteome analysis provides a potential promising approach in disease diagnosis and therapeutic monitoring. However, the large dynamic range of pr oteins, high-abundant proteins, excess of salt and lipid in serum makes the analysis very challenging. Therefore, it is imperative to improve the dissolution of proteins in twodimensional gel electrophoresis (2-DE), and enhance the ability to analyze the proteome of serum under a wide variety of physiological conditions. This study examined the effects of various combinations of depletion highabundant protein, precipitated means, hot SDS treatment and IPG strips with different pH on 2-DE map for mouse serum. Finally the removal of high-abundant proteins in serum by the ProteoExtract TM Albumin Removal column, ethanol precipitation, the heating with 2.5% SDS and 2.3% DTT to denature sample at 95 o C for 3 min, and IEF on pH 4-7 IPG strips (17cm) with 100 µg depleted serum proteins are generally recommended for serum proteome analysis on 2-DE by silver staining, which can effectively improve the resolution and intensity of low-abundant proteins. The optimized conditions help to produce a better reference 2-DE gel of serum samples for following identification potential novel disease markers.