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.
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.
Li
Chen
C
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.
Hong
Yi
Y
Tan
Ye-Xiong
YX
Zhou
Hu
H
Ai
Jian-Hua
JH
Li
Su-Jun
SJ
Zhang
Lei
L
Xia
Qi-Chang
QC
Wu
Jia-Rui
JR
Wang
Hong-Yang
HY
Zeng
Rong
R
D000345:Affinity Labels / Q000032:analysis* / Q000378:metabolism; D006528:Carcinoma, Hepatocellular / Q000737:chemistry / Q000378:metabolism* / Q000473:pathology; D002853:Chromatography, Liquid; D015180:Electrophoresis, Gel, Two-Dimensional; D006801:Humans; D007554:Isotopes / Q000737:chemistry; D007834:Lasers; D008113:Liver Neoplasms / Q000737:chemistry / Q000378:metabolism / Q000473:pathology; D008297:Male; D013058:Mass Spectrometry; D042282:Microdissection; D008875:Middle Aged; D020543:Proteome / Q000032:analysis*; D040901:Proteomics / Q000379:methods*
D000345:Affinity Labels; D007554:Isotopes; D020543:Proteome