Microscale epitope mapping by affinity capillary electrophoresis-mass spectrometry.

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.