Integrated microfluidic system for proteomic analysis consisting of on-line protein digestion, peptides separation and identification

	Integrated microfluidic system for proteomic analysis consisting of on-line protein digestion, peptides separation and identification
	Liang Y(梁玉) ; Tao DY(陶定银) ; Liang Z(梁振) ; Zhang LH(张丽华) ; Zhang YK(张玉奎)
	2011
会议名称	35th international symposium on high performance liquid phase separations and related techniques
会议日期	2010-6-19
会议地点	boston
页码	0-0
通讯作者	张丽华
中文摘要	recently, microfluidic systems have been paid much attention to achieve high throughput proteome analysis. in our recent work, an integrated microfluidic platform, consisting of on-line protein digestion by immobilized enzymatic reactor (imer) prepared in a microchannel, peptides separation and identification by nanorplc-esi-ms/ms, was developed and successfully applied into the analysis of e. coli cell lysate. to achieve rapid on-line protein digestion, a novel monolithic hydrophilic polymer-based imer was prepared within the specified position of a microchannel, by photopolymerization of n-acryloxysuccinimide and poly (ethylene glycol) diacrylate, followed by trypsin immobilization via succinimide functionalities, by which 4μg myoglobin could be digested within 6 min, with sequence coverage as 81.5%. to achieve on-line separation and identification of protein digests, a fused-silica capillary (6-cm length, 75-μm i.d., 190-μm o.d.) with one end pulled to a fine point of ca. 5 μm was packed with c18 particles (5 m, 300å), glued to the outlet channel of imer without dead volume, and coupled with esi-ms/ms directly. sample could be introduced via a pump and a valve (a) connected to the inlet of imer. another microchannel vertical to the outlet of imer was fabricated, and connected to binary pumps via another valve (b). with valve b blocked, proteins were introduced into imer, on-line digested, and trapped on c18 tip. then, with valve a blocked, peptides were separated by the nanorplc column and identified by esi-ms/ms. by such an integrated microfluidic platform, within 85 min, a mixture of bsa, myoglobin and cytochrome c were identified with the average sequence coverage as 51.29%, 60.78% and 57.69%, respectively. in addition, ca. 50 proteins were identified from one 5-min rplc fraction of e. coli cell lysate.
合作状况	墙报
会议主办者	casss
会议录	integrated microfluidic system for proteomic analysis consisting of on-line protein digestion, peptides separation and identification
会议录出版者	待补充
会议录出版地	待补充
学科主题	分析化学
内容类型	会议论文
源URL	[http://159.226.238.44/handle/321008/115952]
专题	大连化学物理研究所_中国科学院大连化学物理研究所
推荐引用方式 GB/T 7714	Liang Y,Tao DY,Liang Z,et al. Integrated microfluidic system for proteomic analysis consisting of on-line protein digestion, peptides separation and identification[C]. 见:35th international symposium on high performance liquid phase separations and related techniques. boston. 2010-6-19.