Picomolar Assay of Native Proteins by Capillary Electrophoresis Precolumn Labeling, Submicellar Separation, and Laser-Induced Fluorescence Detection

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• 作者：Devanand M. Pinto ; Edgar A. Arriaga ; Doug Craig ; Jordanka Angelova ; Neepun Sharma ; Hossein Ahmadzadeh ; Norman J. Dovichi ; and Camille A. Boulet
• 刊名：Analytical Chemistry
• 出版年：1997
• 出版时间：August 1, 1997
• 年：1997
• 卷：69
• 期：15
• 页码：3015 - 3021
• 全文大小：285K
• 年卷期：v.69,no.15(August 1, 1997)
• ISSN：1520-6882

文摘

We report a method for the assay of proteins at concentrations lower than 10^-10 M with as little as 200amol ofprotein. High sensitivity is accomplished byderivatizingthe mages/gifchars/epsilon.gif" BORDER=0 >-amino group of the protein's lysine residues withthe fluorogenic dye 5-furoylquinoline-3-carboxaldehydeand use of a sheath flow cuvette fluorescence detector.Most proteins have a large number of lysine residues;therefore, a large number of fluorescent molecules canbe attached to each protein molecule. In general, precolumn labeling improves sensitivity but degrades resolution due to the inhomogeneity of the reaction productsfrom multiple labeling. However, we demonstrate that,through careful manipulation of the separation and reaction conditions, high sensitivity can be obtained withoutexcessive loss in separation efficiency. Over190 000theoretical plates are obtained for fluorescently labeledovalbumin.