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学科主题分析化学
Integrated Microfluidic System Containing Two Dimensional Separation and MS Identification for Proteomics Analysis
Liang Y(梁玉); Zhang LH(张丽华); Zhang YK(张玉奎); Liang Z(梁振); Dai ZP(戴忠鹏); Liang ZC(梁作成)
会议文集Pittsburgh Conference on Analytical Chemistry and Applied Spectroscopy
会议名称Pittsburgh Conference on Analytical Chemistry and Applied Spectroscopy
会议日期2011-3-13
2011
会议地点亚特兰大
页码1274-0
出版者待补充
出版地待补充
合作性质墙报
部门归属1810
主办者匹兹堡光谱学会
英文摘要Microfluidic systems have been paid much attention to achieve high throughput proteomics analysis, with the advantages of improved portablility, low sample and reagent consumption, accelerated analytical speed, and facile integration. In this paper, the microchip containing SCX monolith and C18 tip was developed and integrated with ESI-MS/MS for two dimensional separation and MS identification of peptides. The SCX monolith in the microchannel was prepared by the photopolymerization of ethylene glycol methacrylate phosphate and bisacrylamide in a trinary porogenic solvent, consisting of dimethylsulfoxide, dodecanol, and N,N-dimethylformamide, which exhibits high dynamic binding capacity and high permeability. The C18 tip was prepared by a fused-silica capillary with a pulled 5-m needle tip, packed with C18 particles (5 m, 300 Å), inset and glued to the outlet of microchannel, coupled with ESI-MS/MS directly, used as the separation column and the chip-to-MS interface. 4 g tryptic digests of membrane proteins extracted from rat liver were loaded on SCX monolith by buffer C (0.1% formic acid water). Then a series of stepwise elution with salt concentration of 250, 350, 500, 750 and 1000 mM CH3COONH4 adjusted by buffer C and buffer D (1000 mM CH3COONH4 at pH 3) was used to elute peptides from SCX monolith onto C18 tip, followed by nano-RPLC-ESI-MS/MS analysis. After database searching using MASCOT with the significance threshold p<0.05, Ions score cut-off 20, and bold red required, 295 proteins were successfully identified with 879 peptides, containing 120 transmembrane proteins, which demonstrated the applicability of such a system for proteomics analysis.; Microfluidic systems have been paid much attention to achieve high throughput proteomics analysis, with the advantages of improved portablility, low sample and reagent consumption, accelerated analytical speed, and facile integration. In this paper, the microchip containing SCX monolith and C18 tip was developed and integrated with ESI-MS/MS for two dimensional separation and MS identification of peptides. The SCX monolith in the microchannel was prepared by the photopolymerization of ethylene glycol methacrylate phosphate and bisacrylamide in a trinary porogenic solvent, consisting of dimethylsulfoxide, dodecanol, and N,N-dimethylformamide, which exhibits high dynamic binding capacity and high permeability. The C18 tip was prepared by a fused-silica capillary with a pulled 5-m needle tip, packed with C18 particles (5 m, 300 Å), inset and glued to the outlet of microchannel, coupled with ESI-MS/MS directly, used as the separation column and the chip-to-MS interface. 4 g tryptic digests of membrane proteins extracted from rat liver were loaded on SCX monolith by buffer C (0.1% formic acid water). Then a series of stepwise elution with salt concentration of 250, 350, 500, 750 and 1000 mM CH3COONH4 adjusted by buffer C and buffer D (1000 mM CH3COONH4 at pH 3) was used to elute peptides from SCX monolith onto C18 tip, followed by nano-RPLC-ESI-MS/MS analysis. After database searching using MASCOT with the significance threshold p<0.05, Ions score cut-off 20, and bold red required, 295 proteins were successfully identified with 879 peptides, containing 120 transmembrane proteins, which demonstrated the applicability of such a system for proteomics analysis.
语种英语
文献类型会议论文
条目标识符http://cas-ir.dicp.ac.cn/handle/321008/116033
专题中国科学院大连化学物理研究所
通讯作者Zhang LH(张丽华)
推荐引用方式
GB/T 7714
Liang Y,Zhang LH,Zhang YK,et al. Integrated Microfluidic System Containing Two Dimensional Separation and MS Identification for Proteomics Analysis[C]. 待补充:待补充,2011:1274-0.
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