DICP OpenIR
Subject Area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)
Source PublicationPittsburgh Conference on Analytical Chemistry and Applied Spectroscopy
Conference NamePittsburgh Conference on Analytical Chemistry and Applied Spectroscopy
Conference Date2011-3-13
2011
Conference Place亚特兰大
Pages1274-0
Publisher待补充
Publication Place待补充
Cooperation Status墙报
Department1810
Funding Organization匹兹堡光谱学会
AbstractMicrofluidic 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.
Language英语
Document Type会议论文
Identifierhttp://cas-ir.dicp.ac.cn/handle/321008/116033
Collection中国科学院大连化学物理研究所
Corresponding AuthorZhang LH(张丽华)
Recommended Citation
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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