AIM： To conduct bhe proteomic analysis of human colorectal carcinoma cell line, SW480 by using two-dimensional electrophoresis （2-DE） and matrix-assisted laser desorption /ionization-time of flight mass spectromet...AIM： To conduct bhe proteomic analysis of human colorectal carcinoma cell line, SW480 by using two-dimensional electrophoresis （2-DE） and matrix-assisted laser desorption /ionization-time of flight mass spectrometry （MALDITOFMS）. METHODS： The total proteins of human colorectal carcinoma cell line, SW480 were separated with 2-DE by using immobilized pH gradient strips and visualized by staining with silver nitrate. The gel images were acquired by scanner and 2-DE analysis software, Image Master 2D Elite. Nineteen distinct protein spots were excised from gel randomly and digested in gel by TPCK-trypsin. Mass analysis of the byptic digest peptides mixture was performed by using MALDI-TOF MS. Peptide mass fingerprints （PMFs） obtained by the MALDI-TOF analysis were used to search NCBI, SWISS-PROT and MSDB databases by using Mascot software. RESULTS： PMF maps of all spots were obtained by MALDI-TOF MS and thirteen proteins were preliminarily identified. CONCLUSION： The methods of analysis and identification of protein spots of tumor cells in 2-DE gel with silver staining by MALDI-TOF MS derived PMF have been established. Protein expression profile of SW480 has been obtained. It is demonstrated that a combination of proteomics and cell culture is a useful approach to comprehend the process of colon carcinogenesis.展开更多
基金Supported by the Natural Science Foundation, Y100-573006Doctoral Foundation of Xi'an Jiaotong University, DFXJTU2002-11
文摘AIM： To conduct bhe proteomic analysis of human colorectal carcinoma cell line, SW480 by using two-dimensional electrophoresis （2-DE） and matrix-assisted laser desorption /ionization-time of flight mass spectrometry （MALDITOFMS）. METHODS： The total proteins of human colorectal carcinoma cell line, SW480 were separated with 2-DE by using immobilized pH gradient strips and visualized by staining with silver nitrate. The gel images were acquired by scanner and 2-DE analysis software, Image Master 2D Elite. Nineteen distinct protein spots were excised from gel randomly and digested in gel by TPCK-trypsin. Mass analysis of the byptic digest peptides mixture was performed by using MALDI-TOF MS. Peptide mass fingerprints （PMFs） obtained by the MALDI-TOF analysis were used to search NCBI, SWISS-PROT and MSDB databases by using Mascot software. RESULTS： PMF maps of all spots were obtained by MALDI-TOF MS and thirteen proteins were preliminarily identified. CONCLUSION： The methods of analysis and identification of protein spots of tumor cells in 2-DE gel with silver staining by MALDI-TOF MS derived PMF have been established. Protein expression profile of SW480 has been obtained. It is demonstrated that a combination of proteomics and cell culture is a useful approach to comprehend the process of colon carcinogenesis.
