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

Fengming Gong, Shufang Liang, Chunmei He, Guobo Shen, Yuhuan Xu, Zhizhong Xu, Zhenling Wang, Xiaohua Wu, Yuquan Wei
2008 Journal of Proteomics & Bioinformatics  
Serum proteome analysis provides a potential promising approach in disease diagnosis and therapeutic monitoring. However, the large dynamic range of proteins, 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
more » ... fects 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.
doi:10.4172/jpb.1000032 fatcat:hfhpcmulwzhkpnvjn5smbtbewa