Nano-flow multidimensional liquid chromatography with electrospray ionization time-of-flight mass spectrometry for proteome analysis of hepatocellular carcinoma |
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Authors: | Yan Wang Jie Zhang Chun-Li Liu Xue Gu Xiang-Min Zhang |
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Institution: | Department of Chemistry, Fudan University, Shanghai 200433, China Research Center of Proteome, Fudan University, Shanghai 200032, China |
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Abstract: | Hepatocellular carcinoma (HCC) is one of the top five cancers with the highest incident of a disease worldwide. To understand the mechanisms of hepatocarcinogenesis, proteomics analysis provides a powerful tool to identify proteins that associate with HCC. We developed a two-step procedure for mapping of HCC proteomics. In the first step, in order to simplify the complexity of proteomics of HCC, the subfractionation of complex protein mixtures in HCC into “subproteomes” is presented based on the solubility of protein. While in the second step an automate comprehensive two-dimensional (2D) separation system, coupling strong cation-exchange (SCX) in the first dimension with capillary reversed-phase chromatography (cRPLC) in the second dimension is developed further to separate and analyze proteins associated with HCC. By using this system, complex sample can be injected, desalted, separated and analyzed in complete automatization. The procedure for proteomics analysis was found to be applied for proteins with great molecular mass (>100 000), small molecular mass (<20 000), highly basic (pI > 9.5) and hydrophobicity, which are not well resolved in 2D-gel electrophoresis. In total 229 proteins were identified by using the described proteomics platform. Among them, several proteins related to the process of carcinogenesis were investigated further. |
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Keywords: | Human hepatocellular carcinoma Two-dimensional liquid chromatography Electrospray ionization mass spectrometer Proteomics Nano-flow |
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