Optimization of 2-dimensional gel electrophoresis for proteomic studies of solid tumor tissue samples

Xu Liang, Jing Rong Wang, Kam Wai V. Wong, Wendy W L HSIAO, Hua Zhou, Zhi Hong JIANG, Kin Ting R. Kam, Liang LIU*

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

6 Citations (Scopus)

Abstract

The method of 2-dimensional gel electrophoresis (2-DE) has been widely used for the proteomic profiling of solid biological samples, however, the analytical conditions have not been optimized. The present study optimized the major conditions of 2-DE for determining the protein contents of solid tumor tissues, through enhancement of the separation efficiency and resolution. Three major analytical conditions of 2-DE analysis, namely protein extraction, focusing time for isoelectric focusing (IEF), and pre-reduction and alkylation prior to IEF, were carefully examined so that the optimal parameters and procedures were achieved. The use of a bead mill for protein extraction resulted in a higher protein yield in a minimal processing time. An optimal focusing time for IEF was established which improved the 2-DE image quality and reproducibility. Furthermore, reduction and alkylation of the protein sample prior to IEF reduced the horizontal streaking caused by oxidation and improved the resolution at the cathode. The optimized 2-DE analysis enabled the detection of 20% more protein spots compared with the previous reported conditions, with higher image quality and reproducibility. Accordingly, the optimized conditions may be used in the 2-DE analysis of tumor tissue samples, by which novel biomarkers of cancerous diseases and molecular targets of drugs are expected to be identified.

Original languageEnglish
Pages (from-to)626-632
Number of pages7
JournalMolecular Medicine Reports
Volume9
Issue number2
DOIs
Publication statusPublished - Feb 2014

Scopus Subject Areas

  • Biochemistry
  • Molecular Medicine
  • Molecular Biology
  • Genetics
  • Oncology
  • Cancer Research

User-Defined Keywords

  • 2-dimensional gel electrophoresis
  • Optimization method
  • Proteomics
  • Tumor

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