Analysis of Human Lung Squamous Carcinoma Cell Line NCI H520 Proteome by Two Dimensional Electrophoresis and MALDI TOF Mass Spectrometry
10.3321/j.issn:1000-467X.2001.06.003
- VernacularTitle:双向凝胶电泳-飞行时间质谱法分析人肺鳞癌细胞NCI-H520蛋白质组成分
- Author:
Quan Xian ZHAN
1
;
Chu Zhu CHEN
;
Jun Yong GUAN
;
Cui LI
;
Mei Chun HE
;
Ping Song LIANG
;
Yun Jin XIE
;
Ping CHEN
Author Information
1. Central South University
- From:Chinese Journal of Cancer
2001;20(6):575-582
- CountryChina
- Language:Chinese
-
Abstract:
Objective：This study was designed to establish and optimize the research methods for proteome,and to analyze the proteome components of human lung squamous carcinoma cell line NCI H520. Methods： A series of methods, including immobilized pH gradient two dimensional polyacrylamide gel electrophoresis(2DE), silver staining, PDQuest 2DE analysis software, peptide mass fingerprint based on matrix assisted laser desorption/ionization time of flying mass spectrometry (MALDI TOF MS) and SWISS PROT database searching, were used to separate and indentify the proteome of human lung squarmous carcinoma cell line NCI H520. Results： The good 2DE pattern including resolution and reproducibility was obtained. After silver staining, the 2DE image analysis by PDQuest 2DE software had detected average of 1146± 116 spots, and 851± 95 spots were matched. The average matching rate was 73.3％ . There had a good reproducibility of spot position in 2DE map, with average deviation in IEF direction of 1.52± 0.22 mm， while in SDS PAGE direction it was 1.97± 0.13 mm. Sixty spots were incised from silver staining gel randomly and digested in gel by TPCKtrypsin. Fifty four peptide mass fingerprints (PMF) maps were obtained by MALDI TOF MS. The typic peptide masses were searched in the SWISS PROT database by PeptIdent software. Forty four proteins were preliminarily identified. Some of them were cell cycle related proteins such as Cyclin H, some were signal transduction related proteins such as mitogen activated protein kinase, protein kinase C and receptor protein tyrosine kinase ERBB 2, some were oncogene related proteins such as Ras related protein RAB 36, etc. Conclusions： The main proteome research system including IPG 2DE, image analysis, MALDI TOF MS derived PMFs and database searching has been established. The data of NCI H520 obtained by above methods will be useful for the establishment of human lung squamous cell proteome database.
