Fast preparation of monolithic immobilized pH gradient column by photopolymerization and photografting techniques for isoelectric focusing separation of proteins
Fast preparation of monolithic immobilized pH gradient column by photopolymerization and photografting techniques for isoelectric focusing separation of proteins
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DOI：
10.1002/elps.201100195
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发表时间：
2011-10-01
期刊：
影响因子：
2.9
通讯作者：
Zhang, Yukui
中科院分区：
文献类型：
--
作者：
Liang, Yu;Zhu, Guijie;Zhang, Yukui
在紫外透明石英毛细管柱中，通过丙烯酰胺和N，N '-亚甲基双丙烯酰胺的光聚合反应5 min，然后将聚焦的两性酚衍生的甲基丙烯酸缩水甘油酯单体光接枝到整体柱上20 min，制备了固定化pH梯度整体柱（M-IPG），缩短了制备时间，提高了形成的pH梯度的稳定性，与我们以前的方法相比。利用制备的M-IPG柱，根据蛋白质的pI实现蛋白质的基线分离。在运行缓冲液中不加入载体两性电解质，分离组分可在低波长下用紫外检测器进行高灵敏度检测。
A new method was developed to prepare monolithic immobilized pH gradient (M-IPG) columns in UV-transparent fused-silica capillaries by the 5-min photopolymerization of acrylamide and N,N'-methylenebisacrylamide, followed by the 20-min photografting of the focused ampholine-derived glycidylmethacrylate monomer on the monolithic matrix, by which the preparation time was reduced, and the stability of the formed pH gradient was improved, compared with our previous methods. Using the prepared M-IPG column, the baseline separation of proteins was achieved according to their pIs. Without carrier ampholytes added in the running buffer, the separated components could be detected with high sensitivity by UV at low wavelength.