Electrophoretic separation of proteins on a microchip with noncovalent, postcolumn labeling.
Electrophoretic separation of proteins on a microchip with noncovalent, postcolumn labeling.
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通过非共价柱后标记在微芯片上电泳分离蛋白质。
DOI：
10.1021/ac000625f
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发表时间：
2000
影响因子：
7.4
通讯作者：
Ramsey,JM
中科院分区：
文献类型：
--
作者：
Liu,Y;Foote,RS;Jacobson,SC;Ramsey,RS;Ramsey,JM
蛋白质通过微芯片毛细管电泳分离，柱后添加荧光染料NanoOrange在芯片上标记，用于激光诱导荧光检测。NanoOrange与疏水蛋白区域非共价结合，形成高荧光复合物。芯片上络合物形成的动力学测量表明，在用于蛋白质分离的条件下，反应速率接近扩散极限。列后标记步骤很少或没有引起带加宽。模型蛋白α-乳白蛋白、β-乳球蛋白A和β-乳球蛋白B的检出下限均<0.5 pg （~ 30 amol）。将其相对荧光和反应速率与其他一些用于蛋白质标记的荧光染料进行了比较。
Proteins were separated by microchip capillary electrophoresis and labeled on-chip by postcolumn addition of a fluorogenic dye, NanoOrange, for detection by laser-induced fluorescence. NanoOrange binds noncovalently with hydrophobic protein regions to form highly fluorescent complexes. Kinetic measurements of complex formation on the microchips suggest that the reaction rate is near the diffusion limit under the conditions used for protein separation. Little or no band broadening is caused by the postcolumn labeling step. Lower limits of detection for model proteins, α-lactalbumin, β-lactoglobulin A, and β-lactoglobulin B, were <0.5 pg (∼30 amol) of injected sample. The relative fluorescence and reaction rates are compared with those of a number of other fluorogenic dyes used for protein labeling.