Abstract
Two-dimensional fluorescence difference gel electrophoresis (2D DIGE) has become a general platform for analysis of various clinical samples such as biofluids and tissues. In comparison to conventional 2-D polyacrylamide gel electrophoresis (2D PAGE), 2D DIGE offers several advantages, such as accuracy and reproducibility between experiments, which facilitate spot-to-spot comparisons. Although whole plasma can be easily obtained, the complexity of plasma samples makes it challenging to analyze samples with good reproducibility. Here, we describe a method for decreasing protein complexity in plasma samples within a narrow pH range by depleting high-abundance plasma proteins. In combination with analysis of differentially expressed spots, trypsin digestion, identification of protein by mass spectrometry, and standard 2D PAGE and DIGE, this method has been optimized for comparison of plasma samples from healthy donors and patients diagnosed with hepatocellular carcinoma.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
References
Paik YK, Kim H, Lee EY et al (2008) Overview and introduction to clinical proteomics. Methods Mol Biol 428:1–31
Hochstrasser DF, Sanchez JC, Appel RD (2002) Proteomics and its trends facing nature’s complexity. Proteomics 2(7):807–812
Cho SY, Lee EY, Kim HY et al (2008) Protein profiling of human plasma samples by two-dimensional electrophoresis. Methods Mol Biol 428:57–75
Cho SY, Lee EY, Lee JS et al (2005) Efficient prefractionation of low--abundance proteins in human plasma and construction of a two-dimensional map. Proteomics 5(13): 3386–3396
Parkin DM, Bray F, Ferlay J et al (2005) Global cancer statistics. 2002. CA Cancer J Clin 55:74–108
Wright LM, Kreikemeie JT, Fimmel CJ (2007) A concise review of serum markers for hepatocellular cancer. Cancer Detect Prev 31: 35–44
Daniele B, Bencivenga A, Megna AS et al (2004) Alpha-fetoprotein and ultrasonography screening for hepatocellular carcinoma. Gastroenterology 127:S108–S112
Sun S, Lee NP, Poon RT et al (2007) Oncoproteomics of hepatocellular carcinoma: from cancer markers’ discovery to functional pathways. Liver Int 27(8):1021–1038
SantamarÃa E, Muñoz J, Fernández--Irigoyen J et al (2007) Toward the discovery of new biomarkers of hepatocellular carcinoma by proteomics. Liver Int 27(2):163–173
Lilley KS, Friedman DB (2004) All about DIGE: quantification technology for differential-display 2D-gel proteomics. Expert Rev Proteomics 1(4):401–409
Park KS, Kim H, Kim NG et al (2002) Proteomic analysis and molecular characterization of tissue ferritin light chain in hepatocellular carcinoma. Hepatology 2002 35(6):1459–1466
Park KS, Cho SY, Kim H et al (2002) Proteomic alterations of the variants of human aldehyde dehydrogenase isozymes correlate with hepatocellular carcinoma. Int J Cancer 97(2):261–265
Lee HJ, Lee EY, Kwon MS et al (2006) Biomarker discovery from the plasma proteome using multidimensional fractionation proteomics. Curr Opin Chem Biol 10(1):42–49
Lee HJ, Na K, Kwon MS et al (2009) Quantitative analysis of phosphopeptides in search of the disease biomarker from the hepatocellular carcinoma specimen. Proteomics 9(12):3395–3408
Jeong SK, Lee EY, Cho JY et al (2010) Data management and functional annotation of the Korean reference plasma proteome. Proteomics 10(6):1250–1255
Jeong SK, Kwon MS, Lee EY et al (2009) BiomarkerDigger: a versatile disease proteome database and analysis platform for the identification of plasma cancer biomarkers. Proteomics 9(14):3729–3740
Na K, Lee EY, Lee HJ et al (2009) Human plasma carboxylesterase 1, a novel serologic biomarker candidate for hepatocellular carcinoma. Proteomics 9(16):3989–3999
Friedman DB, Lilley KS (2008) Optimizing the difference gel electrophoresis (DIGE) technology. Methods Mol Biol 428:93–124
Yan JX, Kett WC, Herbert BR et al (1998) Identification and quantitation of cysteine in proteins separated by gel electrophoresis. J Chromatogr A 813:187–200
Acknowledgments
This study was supported by a grant from the National R&D Program for Cancer Control (1120200 to YKP), National Project for the Personalized Medicine, Ministry for Health and Welfare, Republic of Korea (A111218-11-CP01 to YKP). The authors declare no conflicts of interest. Corresponding author: Young-Ki Paik (paikyk@yonsei.ac.kr).
Author information
Authors and Affiliations
Corresponding author
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2012 Springer Science+Business Media, LLC
About this protocol
Cite this protocol
Na, K., Lee, MJ., Jeong, HJ., Kim, H., Paik, YK. (2012). Differential Gel-Based Proteomic Approach for Cancer Biomarker Discovery Using Human Plasma. In: Cramer, R., Westermeier, R. (eds) Difference Gel Electrophoresis (DIGE). Methods in Molecular Biology, vol 854. Humana Press. https://doi.org/10.1007/978-1-61779-573-2_16
Download citation
DOI: https://doi.org/10.1007/978-1-61779-573-2_16
Published:
Publisher Name: Humana Press
Print ISBN: 978-1-61779-572-5
Online ISBN: 978-1-61779-573-2
eBook Packages: Springer Protocols