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Originally published In Press as doi:10.1074/mcp.M300133-MCP200 on January 15, 2004.
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Molecular & Cellular Proteomics 3:399-409, 2004.
© 2004 by The American Society for Biochemistry and Molecular Biology, Inc.

Research

Accurate Qualitative and Quantitative Proteomic Analysis of Clinical Hepatocellular Carcinoma Using Laser Capture Microdissection Coupled with Isotope-coded Affinity Tag and Two-dimensional Liquid Chromatography Mass Spectrometry*

Chen Li{ddagger}, Yi Hong§, Ye-Xiong Tan§, Hu Zhou{ddagger}, Jian-Hua Ai§, Su-Jun Li{ddagger}, Lei Zhang{ddagger}, Qi-Chang Xia{ddagger}, Jia-Rui Wu{ddagger}, Hong-Yang Wang§,¶ and Rong Zeng{ddagger},¶

From the {ddagger} Research Center for Proteome Analysis, Key Lab of Proteomics, Institute of Biochemistry and Cell Biology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai 200031, China; and § Eastern Hepatobiliary Surgery Hospital, Shanghai 200438, China

Laser capture microdissection (LCM) is a powerful tool that enables the isolation of specific cell types from tissue sections, overcoming the problem of tissue heterogeneity and contamination. This study combined the LCM with isotope-coded affinity tag (ICAT) technology and two-dimensional liquid chromatography to investigate the qualitative and quantitative proteomes of hepatocellular carcinoma (HCC). The effects of three different histochemical stains on tissue sections have been compared, and toluidine blue stain was proved as the most suitable stain for LCM followed by proteomic analysis. The solubilized proteins from microdissected HCC and non-HCC hepatocytes were qualitatively and quantitatively analyzed with two-dimensional liquid chromatography tandem mass spectrometry (2D-LC-MS/MS) alone or coupled with cleavable ICAT labeling technology. A total of 644 proteins were qualitative identified, and 261 proteins were unambiguously quantitated. These results show that the clinical proteomic method using LCM coupled with ICAT and 2D-LC-MS/MS can carry out not only large-scale but also accurate qualitative and quantitative analysis.

To whom correspondence should be addressed: Hong-Yang Wang, Eastern Hepatobiliary Surgery Hospital, No. 225, Changhai Road, Shanghai 200438, China. Tel.: 86-21-25070856; Fax: 86-21-65566851; E-mail: hywangk{at}online.sh.cn; Rong Zeng, Research Center for Proteome Analysis, Institute of Biochemistry and Cell Biology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, 320 YueYang Road, Shanghai 200031, China. Tel.: 86-21-54920170; Fax: 86-21-54920171; E-mail: zr{at}sibs.ac.cn


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