Microscale epitope mapping by affinity capillary electrophoresis-mass spectrometry.
Microscale epitope mapping by affinity capillary electrophoresis-mass spectrometry.
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DOI：
10.1021/ac9700944
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发表时间：
1997-08
影响因子：
7.4
通讯作者：
Y. Lyubarskaya;Y. Dunayevskiy;P. Vouros;B. Karger
中科院分区：
文献类型：
--
作者：
Y. Lyubarskaya;Y. Dunayevskiy;P. Vouros;B. Karger
以β-内啡肽为模型体系，提出了一种基于亲和毛细管电泳-质谱（ACE-MS）的线性表位定位新方法。胰蛋白酶肽在中性涂层毛细管中分离，并通过紫外吸收和电喷雾质谱法进行监测。然后，在注射肽消化物混合物后，注射抗β-内啡肽抗体。与抗体结合的肽被捕获并从其迁移时间中消失。在该消减筛选程序之后，通过ACE-MS程序检查单独合成或分离的免疫反应性肽的结合，以确认表位位于肽上。然后可以制备一系列截短的肽，并通过ACE-MS确定精确的表位。该方法每次运行利用低飞摩尔量的抗体和肽消化物，并且快速且易于自动化。
Using beta-endorphin as a model system, a new microscale solution-based approach for linear epitope mapping based on affinity capillary electrophoresis-mass spectrometry (ACE-MS) is demonstrated. Tryptic peptides are separated in a neutral coated capillary and monitored by ultraviolet absorbance and electrospray mass spectrometry. Then, following injection of the peptide digest mixture, anti-beta-endorphin antibody is injected. The peptide, which binds to the antibody, is captured and disappears from its migration time. Following this subtraction-screening procedure, the binding of the individually synthesized or isolated immunoreactive peptide is examined by the ACE-MS procedure to confirm that the epitope resides on the peptide. A series of truncated peptides can then be made and the precise epitope determined by ACE-MS. The method utilizes low femtomole amounts of antibody and peptide digest per run and is rapid and easily automatable.